Auto/cross-regulation of Hoxb3 expression in posterior hindbrain and spinal cord

The complex and dynamic pattern of Hoxb3 expression in the developing hindbrain and the associated neural crest of mouse embryos is controlled by three separate cis-regulatory elements: element I (region A), element IIIa, and the r5 enhancer (element IVa). We have examined the cis-regulatory element IIIa by transgenic and mutational analysis to determine the upstream trans-acting factors and mechanisms that are involved in controlling the expression of the mouse Hoxb3 gene in the anterior spinal cord and hindbrain up to the r5/r6 boundary, as well as the associated neural crest which migrate to the third and posterior branchial arches and to the gut. By deletion analysis, we have identified the sequence requirements within a 482-bp element III482. Two Hox binding sites are identified in element III482 and we have shown that in vitro both Hoxb3 and Hoxb4 proteins can interact with these Hox binding sites, suggesting that auto/cross-regulation is required for establishing the expression of Hoxb3 in the neural tube domain. Interestingly, we have identified a novel GCCAGGC sequence motif within element III482, which is also required to direct gene expression to a subset of the expression domains except for rhombomere 6 and the associated neural crest migrating to the third and posterior branchial arches. Element III482 can direct a higher level of reporter gene expression in r6, which led us to investigate whether kreisler is involved in regulating Hoxb3 expression in r6 through this element. However, our transgenic and mutational analysis has demonstrated that, although kreisler binding sites are present, they are not required for the establishment or maintenance of reporter gene expression in r6. Our results have provided evidence that the expression of Hoxb3 in the neural tube up to the r5/r6 boundary is auto/cross-regulated by Hox genes and expression of Hoxb3 in r6 does not require kreisler.     

Infectious disease survey of lesser prairie chickens in north Texas

Lesser prairie chicken (Tympanuchus pallidicinctus) abundance, like that of most grassland birds, has declined rangewide for decades. Although habitat loss and degradation are likely ultimate causes for this decline, infectious agents, particularly microparasites, could be proximate contributors. No surveys of pathogenic bacteria or viruses have been published for this species. We surveyed 24 free-living lesser prairie chickens from Hemphill County, Texas (USA), for evidence of exposure to Salmonella typhimurium, S. pullorum, Mycoplasma gallisepticum, M. synoviae, Chlamydophila psittaci, and the avian influenza, Newcastle disease, infectious bronchitis, and reticuloendotheliosis viruses. Two of 18, and eight of 17 samples were seropositive for the Massachusetts and Arkansas serotypes of infectious bronchitis virus, respectively. Five of the eight positive individuals were juveniles, two of which were seropositive for both serotypes. All other serologic and genetic tests were negative. Because the ecological significance of these results is unknown, the pathogenesis, transmission, and/or population-level influences of infectious bronchitis and related avian coronaviruses for lesser prairie chickens deserves further study.     

The mitochondrial presequence translocase: an essential role of Tim50 in directing preproteins to the import channel

Mitochondrial proteins with N-terminal targeting signals are transported across the inner membrane via the presequence translocase, which consists of membrane-integrated channel proteins and the matrix Hsp70 import motor. It has not been known how preproteins are directed to the import channel. We have identified the essential protein Tim50, which exposes its major domain to the intermembrane space. Tim50 interacts with preproteins in transit and directs them to the channel protein Tim23. Inactivation of Tim50 strongly inhibits the import of preproteins with a classical matrix-targeting signal, while preproteins carrying an additional inner membrane-sorting signal do not strictly depend on Tim50. Thus, Tim50 is crucial for guiding the precursors of matrix proteins to their insertion site in the inner membrane.     

The Salton Sea as critical habitat to migratory and resident waterbirds

Concern about the Salton Sea ecosystem, based on potential impacts of increasing salinity, contaminants, disease outbreaks, and large die-offs of birds, is heightened because of tremendous prior loss and degradation of wetland habitat in western North America. In 1999, we used a variety of survey methods to describe patterns of abundance of birds at the Salton Sea and in adjacent habitats. Our results further documented the great importance of the Salton Sea within the Pacific Flyway to wintering, migratory, and breeding waterbirds. Exclusive of Eared Grebes, we estimated about 187000 individual waterbirds at the Salton Sea in January, 88000 in April, 170000 in August, and 261000 in November. Additional surveys of Eared Grebes in November and December suggested the total population of all waterbirds was about 434000 to 583000 in those months, respectively. We also documented breeding by about 14000 pairs of colonial waterbirds. Waterbirds were particularly concentrated along the northern, southwestern, southern, and southeastern shorelines and river deltas. By contrast, some species of wading birds (Cattle Egret, White-faced Ibis, Sandhill Crane) and shorebirds (Mountain Plover, Whimbrel, Long-billed Curlew) were much more numerous in agricultural fields of the Imperial Valley than in wetland habitats at the Sea. Various studies indicate the Salton Sea is of regional or national importance to pelicans and cormorants, wading birds, waterfowl, shorebirds, and gulls and terns. Important taxa are the Eared Grebe, American White Pelican, Double-crested Cormorant, Cattle Egret, White-faced Ibis, Ruddy Duck, Yuma Clapper Rail, Snowy Plover, Mountain Plover, Gull-billed, Caspian, and Black terns, and Black Skimmer. Proposed restoration projects should be carefully assessed to ensure they do not have unintended impacts and are not placed where large numbers of breeding, roosting, or foraging birds concentrate. Similarly, plans to enhance opportunities for recreation or commerce at the Sea should aim to avoid or minimize disturbance to birds. Future research should focus on filling gaps in knowledge needed to effectively conserve birds at the Salton Sea.     

Differential control of vesicle priming and short-term plasticity by Munc13 isoforms

Presynaptic short-term plasticity is an important adaptive mechanism regulating synaptic transmitter release at varying action potential frequencies. However, the underlying molecular mechanisms are unknown. We examined genetically defined and functionally unique axonal subpopulations of synapses in excitatory hippocampal neurons that utilize either Munc13-1 or Munc13-2 as synaptic vesicle priming factor. In contrast to Munc13-1-dependent synapses, Munc13-2-driven synapses show pronounced and transient augmentation of synaptic amplitudes following high-frequency stimulation. This augmentation is caused by a Ca(2+)-dependent increase in release probability and releasable vesicle pool size, and requires phospholipase C activity. Thus, differential expression of Munc13 isoforms at individual synapses represents a general mechanism that controls short-term plasticity and contributes to the heterogeneity of synaptic information coding.     

Efficacy of the biocontrol yeast Pichia anomala during long-term storage of moist feed grain under different oxygen and carbon dioxide regimens

The yeast Pichia anomala inhibits the spoilage mold Penicillium roqueforti in laboratory experiments with high-moisture wheat in malfunctioning airtight storage. The ability of P. anomala to prevent mold growth during 14 months of grain storage was evaluated in outdoor silos with different air permeabilities. Freshly harvested wheat in 160-kg portions was inoculated with 10(2) colony-forming units (cfu) g(-1) P. roqueforti, alone or together with 10(4) cfu g(-1) P. anomala. During the first month P. anomala increased to about 10(6) cfu g(-1) in the treated silos to reach 10(7) cfu g (-1) after 9 months. Naturally occurring P. anomala in the untreated silos increased from 10(2) to about 10(3) cfu g(-1) during the first month and reached the same level as the treated silos after 9 months. Oxygen levels were reduced below the detection limit within 1 day, while carbon dioxide levels increased to 80-90% during the first month. P. roqueforti did not grow in wheat treated with P. anomala, regardless of silo permeability, but had increased to 10(5) cfu g(-1) in the untreated silos after 14 months of storage.     

Dynamic effects of predators on cyclic voles: field experimentation and model extrapolation

Mechanisms generating the well-known 3-5 year cyclic fluctuations in densities of northern small rodents (voles and lemmings) have remained an ecological puzzle for decades. The hypothesis that these fluctuations are caused by delayed density-dependent impacts of predators was tested by replicated field experimentation in western Finland. We reduced densities of all main mammalian and avian predators through a 3 year vole cycle and compared vole abundances between four reduction and four control areas (each 2.5-3 km(2)). The reduction of predator densities increased the autumn density of voles fourfold in the low phase, accelerated the increase twofold, increased the autumn density of voles twofold in the peak phase, and retarded the initiation of decline of the vole cycle. Extrapolating these experimental results to their expected long-term dynamic effects through a demographic model produces changes from regular multiannual cycles to annual fluctuations with declining densities of specialist predators. This supports the findings of the field experiment and is in agreement with the predation hypothesis. We conclude that predators may indeed generate the cyclic population fluctuations of voles observed in northern Europe.     

Overexpression of the Shb SH2 domain-protein in insulin-producing cells leads to altered signaling through the IRS-1 and IRS-2 proteins

BACKGROUND: Overexpression of the Src homology 2 domain protein Shb in beta-cells of transgenic mice has been shown to promote an increased beta-cell mass. To investigate the mechanisms by which Shb controls the beta-cell mass, we have presently studied the effects of Shb overexpression on the IRS-1-induced signaling pathway in mouse islet beta-cells and in insulin-producing RINm5F cells and correlated these effects to growth and death patterns. MATERIALS AND METHODS: Shb overexpression was achieved in RINm5F cells by selection of stable clones or by FACS purification of transiently transfected cells. For Shb overexpression in primary mouse islet cells, a Shb-transgene mouse was used. Cell proliferation and death rates were determined using flow cytometry. Serum-, insulin-, and IGF-1-stimulated signaling events were studied by immunoblot, immunoprecipitation, and in vitro kinase procedures. RESULTS: Transient Shb overexpression in RINm5F cells resulted in increased proliferation. Both Shb-overexpressing RINm5F cells and islet cells from transgenic mice (islet Shb) exhibited increased basal tyrosine phosphorylation of IRS-1. Shb overexpression resulted also in the assembly and activation of a multiunit complex consisting of at least Shb, IRS-1, IRS-2, FAK, and PI3K. Consequently, the phosphorylation of Akt was enhanced under basal conditions in Shb overexpressing cells. Finally, Shb overexpression did not affect insulin-induced phosphorylation of the PI3K-antagonist PTEN. CONCLUSION: It is concluded that the Shb-induced alterations in the IRS-1/PI3K/Akt pathway may be relevant to the understanding of growth and death patterns of insulin-producing cells.     

Segregation of saturated chain lipids in pulmonary surfactant films and bilayers

The physical properties of organized system (bilayers and monolayers at the air water interface) composed of bovine lipid extract surfactant (BLES) were studied using correlated experimental techniques. 6-Dodecanoyl-2-dimethylamino-naphthalene (LAURDAN)-labeled giant unilamelar vesicles (mean diameter approximately 30 microm) composed of BLES were observed at different temperatures using two-photon fluorescence microscopy. As the temperature was decreased, dark domains (gel-like) appeared at physiological temperature (37 degrees C) on the surface of BLES giant unilamelar vesicles. The LAURDAN two-photon fluorescent images show that the gel-like domains span the lipid bilayer. Quantitative analysis of the LAURDAN generalized polarization function suggests the presence of a gel/fluid phase coexistence between 37 degrees C to 20 degrees C with low compositional and energetic differences between the coexisting phases. Interestingly, the microscopic scenario of the phase coexistence observed below 20 degrees C shows different domain's shape compared with that observed between 37 degrees C to 20 degrees C, suggesting the coexistence of two ordered but differently organized lipid phases on the bilayer. Epifluorescence microscopy studies of BLES monomolecular films doped with small amounts of fluorescent lipids showed the appearance and growth of dark domains (liquid condensed) dispersed in a fluorescent phase (liquid expanded) with shapes and sizes similar to those observed in BLES giant unilamelar vesicles. Our study suggests that bovine surfactant lipids can organize into discrete phases in monolayers or bilayers with equivalent temperature dependencies and may occur at physiological temperatures and surface pressures equivalent to those at the lung interface.